Novel single-tube enzymatic fragmentation technology simplifies library preparation, minimizing sample loss
New England Biolabs (NEB®) today announced the release of the NEBNext Ultra II FS DNA Library Prep Kit for next generation sequencing (NGS). This new kit employs a novel enzymatic fragmentation system that combines fragmentation, end repair and dA-tailing in a single tube, and utilizes a single protocol regardless of input amount or GC content.
"As applications for NGS continue to expand and sequencing instrument capacity continues to grow, laboratories must resolve the challenges of quickly preparing high-quality libraries from ever-decreasing input amounts in a scalable and user-friendly manner," said Fiona Stewart, NEBNext Portfolio Manager at NEB.
"With the end user in mind, we've developed a new fragmentation technology that doesn't require the conventional clean-up steps that can result in sample loss. The method is also extremely robust – the same protocol is used for all input DNA amounts and GC contents, and with input DNA in water, TE or Tris. Fragmentation is further integrated into the library prep workflow by combining fragmentation, end repair and dA-tailing reagents into a single mix, enabling subsequent adaptor ligation without a clean-up step. The combination of these high reaction efficiencies and reduction in sample losses increases library yields and enables use of a wide range of input amounts, from 100 pg to 0.5 µg of eukaryotic DNA," said Stewart. "Critically, the quality of the FS libraries is high, with excellent sequencing metrics and GC coverage uniformity, including with PCR-free libraries."
Early access users report that the NEBNext Ultra II FS DNA Kit addresses challenges in DNA fragmentation in a variety of laboratories working at different levels of throughput, including larger genomics and genetic research institutions. Peter Ellis, Senior Staff Scientist at the Wellcome Trust Sanger Institute stated, "Our institute currently processes thousands of DNA samples each month via its core DNA sequencing library construction pipelines. However, a recent requirement to generate high quality whole genome and targeted sequencing data from biopsy material led us to develop and implement a novel workflow enabled by NEBNext Ultra II FS. Our new automated workflow, coupled with the high efficiency of the NEBNext Ultra II FS reagent, is allowing us to routinely generate deep sequence data from as few as 100-1000 human cells."
Dr Luiza Moore, Clinical Research Fellow in the Cancer, Ageing and Somatic Mutation Programme at the Wellcome Trust Sanger Institute added, "This recently developed low input DNA approach that utilises NEBNext Ultra II FS has allowed us to infer genomes derived from small populations of cells of specific types and anatomical locations. Importantly, the method circumvents prior whole genome amplification and associated bias and artefacts."
The NEBNext Ultra II FS DNA Kit has a streamlined workflow and library preparation can be completed within 2.5 hours, with less than 15 minutes of hands-on time. High-efficiency Ultra II ligation reagents are provided for adaptor ligation, and while the kit is compatible with PCR-free workflows with inputs as low as 50 ng of eukaryotic DNA, the NEBNext Q5® Ultra II PCR Master Mix is provided for amplification of libraries when necessary. This kit is also available with the option of SPRISelect® beads for size-selection and clean-up steps.